title: [Construction and identification of a recombinant replication-defective adenovirus vector encoding p53AIP1 and its expression in human HeLa cells].
abstract: OBJECTIVE
  To construct a recombinant replication-defective adenovirus vector carrying p53AIP1 (p53-regulated apoptosis-inducing protein 1) gene and observe its expression in human HeLa cells.
  
  
  METHODS
  Specific primers were designed according to p53AIP1 gene sequence and inserted specific enzyme cutting sites. P53AIP1 gene was amplified by PCR and cloned into the adenovirus shuttle plasmid pDC316 to construct the recombinant vector pDC316-p53AIP1, which was co-transfected with helper plasmid pBHGloxδE1, 3Cre into HEK293 cells by Lipofectamine(TM); 2000. The recombinant replication-defective adenovirus (Ad-p53AIP1) was generated by means of homologous recombination of the two plasmids in HEK293 cells with the Cre-loxP recombinase system and harvested after 12 days. Ad-p53AIP1 and Ad-null were respectively transfected into HeLa cells at MOI=100. Then the expression of p53AIP1 gene was detected by Western blotting.
  
  
  RESULTS
  pDC316-p53AIP1 was constructed successfully. The new constructed vector was confirmed by PCR analysis, double enzyme digestion and DNA sequencing. The results were in conformity with the expected. Western blotting demonstrated that the target p53AIP1 proteins were effectively expressed in the transfected HeLa cells.
  
  
  CONCLUSIONS
  The recombinant adenovirus vector of Ad-p53AIP1 was successfully established, and it was proved to have a strong ability of infectivity.
authors.affiliation: Department of Urology, General Hospital of PLA, Beijing 100853, China.
authors.forename: Xiaoliang
authors.initials: X
authors.lastname: Lin
authors.forename: Yunbo
authors.initials: Y
authors.lastname: Jiang
authors.forename: Qian
authors.initials: Q
authors.lastname: Zhao
authors.forename: Xiaotao
authors.initials: X
authors.lastname: Yin
authors.forename: Renli
authors.initials: R
authors.lastname: Tian
authors.forename: Yunqi
authors.initials: Y
authors.lastname: Li
authors.forename: Yuanji
authors.initials: Y
authors.lastname: Xu
authors.forename: Jinqi
authors.initials: J
authors.lastname: Yan
authors.forename: Wei
authors.initials: W
authors.lastname: Zhang
authors.forename: Jiangping
authors.initials: J
authors.lastname: Gao
authors.forename: Jiyun
authors.initials: J
authors.lastname: Yu
mesh_terms: D000256:Adenoviridae / Q000235:genetics*; D051017:Apoptosis Regulatory Proteins / Q000235:genetics*; D015153:Blotting, Western; D003673:Defective Viruses / Q000235:genetics*; D005822:Genetic Vectors; D006367:HeLa Cells; D006801:Humans; D016133:Polymerase Chain Reaction; D011995:Recombination, Genetic; D014162:Transfection
chemical_list: D051017:Apoptosis Regulatory Proteins; C414742:P53AIP1 protein, human